This research project focuses on the urokinase receptor (uPAR), a multifunctional GPI-anchored membrane protein that has been implicated in breast cancer progression. Our long-term goal is to understand mechanisms by which uPAR regulates cancer cell physiology. uPAR binds at least two ligands: the plasminogen activator, urokinase-type plasminogen activator (uPA), and the provisional extracellular matrix protein, vitronectin. Within the plasma membrane, uPAR laterally associates with integrins, G-protein coupled receptors, and receptor-tyrosine kinases (RTKs). These "co-receptor" interactions result in the formation of a dynamic, multiprotein signaling receptor complex (MSRC). uPA and vitronectin activate uPAR-dependent cell signaling; however, the pathways are distinct. uPA-binding activates ERK/MAP kinase whereas vitronectin activates Rad. Other signaling proteins, such as STAT-5b, may be recruited downstream of uPAR depending on the co-receptors that associate with uPAR in the MSRC. Co-receptors also determine the nature of the cellular response, such as whether uPA is mitogenic. In this application, four specific aims are proposed. In Aim 1, we propose to elucidate the pathway that leads from uPAR to Rac1 and determine the mechanism by which ligation of a single receptor (uPAR) with two distinct ligands (uPA or vitronectin) causes completely distinct cell signaling responses. Aim 2 builds on preliminary data in which we have demonstrated that uPAR over-expression induces epithelial-mesenchymal transformation (EMT), a well known process associated with cancer progression in which epithelial cells adopt a fibroblast-like morphology, lose cell-cell contacts, demonstrate increased vimentin expression and decreased E-cadherin expression. Experiments planned in Aim 2 will determine the mechanism by which uPAR induces EMT. In Aim 3, we will test the hypothesis that uPAR-dependent cell signaling regulates the response of cancer cells not only to uPA but also to growth factors such as EGF and PDGF. Understanding crosstalk between uPAR and RTKs is a critical objective. Finally, in Aim 4, we propose studies to determine whether uPAR-dependent cell signaling is involved in cancer development and progression in vivo. Xenograft and spontaneous neoplasia mouse-model systems will be applied. These studies will elucidate the role of uPAR in cancer and provide critical information regarding how uPAR may be targeted for novel cancer therapeutics design. [unreadable] [unreadable] [unreadable]